Role of Myosin Va in the Plasticity of the Vertebrate Neuromuscular Junction In Vivo

Abstract

BackgroundMyosin Va is a motor protein involved in vesicular transport and its absence leads to movement disorders in humans (Griscelli and Elejalde syndromes) and rodents (e.g. dilute lethal phenotype in mice). We examined the role of myosin Va in the postsynaptic plasticity of the vertebrate neuromuscular junction (NMJ).Methodology/Principal Findings Dilute lethal mice showed a good correlation between the propensity for seizures, and fragmentation and size reduction of NMJs. In an aneural C2C12 myoblast cell culture, expression of a dominant-negative fragment of myosin Va led to the accumulation of punctate structures containing the NMJ marker protein, rapsyn-GFP, in perinuclear clusters. In mouse hindlimb muscle, endogenous myosin Va co-precipitated with surface-exposed or internalised acetylcholine receptors and was markedly enriched in close proximity to the NMJ upon immunofluorescence. In vivo microscopy of exogenous full length myosin Va as well as a cargo-binding fragment of myosin Va showed localisation to the NMJ in wildtype mouse muscles. Furthermore, local interference with myosin Va function in live wildtype mouse muscles led to fragmentation and size reduction of NMJs, exclusion of rapsyn-GFP from NMJs, reduced persistence of acetylcholine receptors in NMJs and an increased amount of punctate structures bearing internalised NMJ proteins.Conclusions/SignificanceIn summary, our data show a crucial role of myosin Va for the plasticity of live vertebrate neuromuscular junctions and suggest its involvement in the recycling of internalised acetylcholine receptors back to the postsynaptic membrane.

Myosin Va cooperates with PKA RIα to mediate maintenance of the endplate in vivo

“…For AMPA receptors, either myosin Va (2) or myosin Vb (1, 3) may be important. We have found myosin Va to be crucial for the maintenance of the neuromuscular synapse, apparently by facilitating recycling of AChRs (4). In extension to this concept, we now present in vivo data showing that for this function, the cooperation of myosin Va with PKA RIα is necessary.…”

“…cAMP | muscle | neuromuscular junction | microdomain | synaptic plasticity R ecently, myosin V motor proteins were found to be crucial for the plasticity of central and peripheral synapses by facilitating, respectively, the recycling of AMPA (1-3) and acetylcholine receptors (AChRs) (4). The neuromuscular junction (NMJ) is the synapse between motor neurons and skeletal muscle fibers.…”

“…Vesicular transport is employed for this purpose, using different routes of exocytosis, endocytosis, and recycling (7)(8)(9). We identified myosin Va as the first motor protein involved in AChR trafficking and have found it to facilitate its recycling (4). Down-regulation of myosin Va led to a reduced t 1/2 of AChRs (4) similar to pathological conditions, such as denervation (6,(10)(11)(12)(13) or dystrophy (14).…”

Role of autophagy, SQSTM1, SH3GLB1, and TRIM63 in the turnover of nicotinic acetylcholine receptors

“…Transfections and sciatic denervations were as previously described. 34,55 Loss of mouse skeletal muscle mass upon denervation was previously measured in other studies and accounted for about 30% (tibialis anterior muscle) and 56% (quadriceps muscle) after 6 19 and 14 d of denervation, 28 respectively. Although muscle weights were not taken in the present study, we determined fiber diameters for all measured fibers from in vivo confocal images.…”

“…Our previous extensive work on the mechanisms underlying CHRN recycling 25,34,35 and CHRN degradation identified an interaction between CHRN and the E3 ligase TRIM63. 28 Furthermore, the previously reported direct interaction between TRIM63 and SQSTM1 29 prompted us to speculate that TRIM63 may induce selective autophagy as a principal route for CHRN removal.…”